Podcast
Questions and Answers
What is the primary use of Coomassie Brilliant Blue in protein analysis?
What is the primary use of Coomassie Brilliant Blue in protein analysis?
Which technique allows for the identification of a specific protein through antibody binding?
Which technique allows for the identification of a specific protein through antibody binding?
What does SDS-PAGE predominantly separate proteins based on?
What does SDS-PAGE predominantly separate proteins based on?
Why is a membrane blocked during a Western Blot procedure?
Why is a membrane blocked during a Western Blot procedure?
Signup and view all the answers
What is a key difference between native PAGE and SDS-PAGE?
What is a key difference between native PAGE and SDS-PAGE?
Signup and view all the answers
What role does EDTA play in the process described?
What role does EDTA play in the process described?
Signup and view all the answers
What would likely happen if EDTA is omitted from the solution?
What would likely happen if EDTA is omitted from the solution?
Signup and view all the answers
What is the reason for using glucose in the lysis solution?
What is the reason for using glucose in the lysis solution?
Signup and view all the answers
Why is a buffer with alkaline pH (Tris pH=8) used in the process?
Why is a buffer with alkaline pH (Tris pH=8) used in the process?
Signup and view all the answers
What is the function of Sodium Acetate (pH=5.5) in the lysis solution?
What is the function of Sodium Acetate (pH=5.5) in the lysis solution?
Signup and view all the answers
What is the primary function of chlorophylls and carotenoids in the reaction center complex?
What is the primary function of chlorophylls and carotenoids in the reaction center complex?
Signup and view all the answers
What process describes the formation of ATP in chloroplasts when light is present?
What process describes the formation of ATP in chloroplasts when light is present?
Signup and view all the answers
Which molecule is reduced to NADPH during the process involving water and light energy?
Which molecule is reduced to NADPH during the process involving water and light energy?
Signup and view all the answers
Where in the chloroplasts are the photosystems primarily located?
Where in the chloroplasts are the photosystems primarily located?
Signup and view all the answers
What role do protons play in ATP generation within chloroplasts?
What role do protons play in ATP generation within chloroplasts?
Signup and view all the answers
Which of the following is NOT a product of the light-dependent reactions of photosynthesis?
Which of the following is NOT a product of the light-dependent reactions of photosynthesis?
Signup and view all the answers
What is the final electron acceptor in the photosynthetic electron transport chain?
What is the final electron acceptor in the photosynthetic electron transport chain?
Signup and view all the answers
Which component of the chloroplasts is responsible for proton pumping during ATP generation?
Which component of the chloroplasts is responsible for proton pumping during ATP generation?
Signup and view all the answers
What is the formula used to make solutions from stock concentrations?
What is the formula used to make solutions from stock concentrations?
Signup and view all the answers
How is a 1% (weight/volume) solution generally defined?
How is a 1% (weight/volume) solution generally defined?
Signup and view all the answers
What is the molarity of a solution that contains 0.5 moles of solute in 2 liters of solution?
What is the molarity of a solution that contains 0.5 moles of solute in 2 liters of solution?
Signup and view all the answers
If you need to prepare 240 mL of a 1X solution from a 15X stock solution, how much of the stock solution do you need to use?
If you need to prepare 240 mL of a 1X solution from a 15X stock solution, how much of the stock solution do you need to use?
Signup and view all the answers
How many particles are there in 1 mole of a substance, according to Avogadro's number?
How many particles are there in 1 mole of a substance, according to Avogadro's number?
Signup and view all the answers
What is the molecular weight (MW) of EDTA as given in the content?
What is the molecular weight (MW) of EDTA as given in the content?
Signup and view all the answers
When creating a diluent for a concentration, what does a 2X solution mean?
When creating a diluent for a concentration, what does a 2X solution mean?
Signup and view all the answers
If the formulation indicates to prepare 150 mL of a 25 mM solution of EDTA, how much EDTA is required?
If the formulation indicates to prepare 150 mL of a 25 mM solution of EDTA, how much EDTA is required?
Signup and view all the answers
Which type of DNA fragment runs faster during agarose gel electrophoresis?
Which type of DNA fragment runs faster during agarose gel electrophoresis?
Signup and view all the answers
What is the role of Ethidium Bromide in gel electrophoresis?
What is the role of Ethidium Bromide in gel electrophoresis?
Signup and view all the answers
Why is GelGreen considered safer than Ethidium Bromide?
Why is GelGreen considered safer than Ethidium Bromide?
Signup and view all the answers
What type of enzyme is responsible for cleaving terminal residues from linear DNA?
What type of enzyme is responsible for cleaving terminal residues from linear DNA?
Signup and view all the answers
What is a characteristic of an isoschizomer?
What is a characteristic of an isoschizomer?
Signup and view all the answers
What issue can result in off-site cleavage during a restriction digest?
What issue can result in off-site cleavage during a restriction digest?
Signup and view all the answers
If plasmid X is 2000 bp and is digested with Enzyme 1 at the 500 bp site, what is the size of the resulting fragment?
If plasmid X is 2000 bp and is digested with Enzyme 1 at the 500 bp site, what is the size of the resulting fragment?
Signup and view all the answers
How many fragments would result when plasmid X is digested with both Enzyme 1 and Enzyme 2?
How many fragments would result when plasmid X is digested with both Enzyme 1 and Enzyme 2?
Signup and view all the answers
What major difference exists between endonuclease and exonuclease enzymes?
What major difference exists between endonuclease and exonuclease enzymes?
Signup and view all the answers
What does it indicate if an enzyme is described as a 'neoschizomer'?
What does it indicate if an enzyme is described as a 'neoschizomer'?
Signup and view all the answers
What is the primary function of BamHI in the context provided?
What is the primary function of BamHI in the context provided?
Signup and view all the answers
What does the notation '6538 bp' represent in the content?
What does the notation '6538 bp' represent in the content?
Signup and view all the answers
If BamHI cuts a fragment of 435 bp, what would be the approximate size of the remaining fragment after the cut?
If BamHI cuts a fragment of 435 bp, what would be the approximate size of the remaining fragment after the cut?
Signup and view all the answers
What does the term 'star activity' refer to in the context given?
What does the term 'star activity' refer to in the context given?
Signup and view all the answers
What is the significance of the 'polyacrylamide gels' mentioned?
What is the significance of the 'polyacrylamide gels' mentioned?
Signup and view all the answers
Which of the following is a potential issue when using restriction enzymes like BamHI?
Which of the following is a potential issue when using restriction enzymes like BamHI?
Signup and view all the answers
What does 'linearizing the plasmid' accomplish in biotechnology?
What does 'linearizing the plasmid' accomplish in biotechnology?
Signup and view all the answers
What can result from running an improper gel during DNA analysis?
What can result from running an improper gel during DNA analysis?
Signup and view all the answers
When three band fragments are formed during a restriction digest, what does this typically indicate?
When three band fragments are formed during a restriction digest, what does this typically indicate?
Signup and view all the answers
What role does the size marker play in gel electrophoresis?
What role does the size marker play in gel electrophoresis?
Signup and view all the answers
Study Notes
Final Exam Review - BCH IS 3201
- Topics needing review include lab math, plasmid digests, calculating fragment size, microarray slides, and using Bragg's law.
Lab Math
- Making solutions from stock using c1V1=c2V2
- % solutions (w/v and v/v)
- Dilutions (1X, 2X, etc.)
- Molarity calculations
- Formula: (Formula weight) x (Volume desired) x (Molarity desired) = g needed
- 1 mole = 6.02 x 10^23 particles
Practice Problems
- Preparing 1L of 0.5M glucose solution (MW 180.2 g/mol)
- Preparing 1L of 100 mM glucose solution (MW 180.2 g/mol)
- Preparing 10mL of 1M glycine solution (MW 75.07 g/mol)
- Preparing 10mL of 0.5M glucose solution (MW 180.2 g/mol)
- Calculating final concentration of 1M glycine in 10mL with 9mL water
- Calculating concentrations of 10mg glucose / 100mL water and 100mL of 2% alanine (89.09 g/mol)
- Making 200mL of 20mM phosphate buffer from 0.5M stock
- Making 1L of 50% (w/v) glucose solution (MW 180.2 g/mol)
Pipetting
- Minimum range on a pipettor is determined as 10% of the maximum range
- Pipettor ranges (ul)
- P10: 0-10 (1-10)
- P20: 0-20 (2-20)
- P200: 0-200 (20-200)
- P1000: 0-1000 (100-1000)
Plasmid Purification - Reaction Components
- EDTA chelates divalent metal cations—preventing DNAse degradation of plasmid DNA
- Glucose prevents instant cell lysis by osmotic shock
- Alkaline pH (Tris pH 8) reduces electrostatic interactions between DNA and scaffolding proteins
Plasmid Purification
- Examining why plasmid DNA renatures and chromosomal DNA does not: plasmid DNA is usually smaller/topologically intertwined than chromosomal DNA
- Examining the use of 90% ethanol as a DNA precipitant
- Discussing the purpose of 70% ethanol.
Agarose Gel Electrophoresis
- Smaller DNA fragments move faster in agarose gel electrophoresis
- Ethidium Bromide and GelGreen: fluorescent dyes intercalating between base pairs for visualization under UV light
- GelGreen is safer than EtBr
Restriction Enzymes
- Restriction enzymes cleave terminal or internal residues from linear DNA, recognizing specific DNA sequences and cleaving at the same or different sites
- Different conditions can cause off-site cleavage (star activity).
Restriction Digest
- Enzyme 1 cleaves at 500 bp
- Enzyme 2 cleaves at 1000bp site
Question - Digesting plasmid Q with Baml and Hindl
- Lane 3: Incomplete digest bands
- Lane 4: Incomplete digest + star activity
Plasmid Digest - Practice
- pBLU plasmid with BamHI - 5437bp fragment
- pBLU plasmid with XmnI and NdeI - different sizes of fragments may be possible.
Question - Digesting Lambda Phage DNA
- Digest with BamHl: various fragment sizes will be created, totalizing 48,502 base pairs
- Digest without completing: various additional fragments, some with variable, incomplete digest bands
Question - Cutting pUC19 with Restriction Enzymes
- How many bands will result from digestion with specific restriction enzymes on a gel
Polyacrylamide Gels
- Resolve proteins or DNA fragments with similar sizes
- Polyacrylamide gels are created with crosslinked acrylamide and bisacryamide
- Native gels separate proteins by size, shape, and charge
- SDS-PAGE separates proteins based on molecular weight.
Native Gel System
- Non-denaturing - protein maintains its natural structure
- Migration depends on charge and size
- Proteins stable post-extraction
SDS Gel System
- Denaturing and reducing - protein unfolds and loses its original form
- Migration only depends on mass (molecular weight)
Western Blot
- Detecting the protein of interest using BLOTTO (milk protein) to block the membrane and antibodies to bind to the target protein
- Identifying specific targets
Polyacrylamide Gel Electrophoresis (PAGE)
- SDS (detergent) coats proteins with uniform negative charge; disrupts protein native structure, and allows migration based on size
- B-mercaptoethanol helps reduce disulfide bridges
Kinetics
- enzyme speed-up rates
- reaction rate (v0)
- substrate concentration ([S])
- Michaelis-Menten equation: V0 = (Vmax * [S])/(Km + [S])
- Vmax: maximum initial reaction rate
- Km: Michaelis constant, substrate concentration at which v = Vmax/2.
Double Reciprocal Plot (Lineweaver-Burk Plot)
- To determine Vmax and Km more precisely.
- The slope = Km/Vmax
- Y-intercept = 1/Vmax
- X-intercept = -1/Km
Types of Inhibition Kinetics
- Competitive, mixed, and uncompetitive inhibition affects both Km and Vmax in different ways.
Additional Questions
- Determining values and interpretations of data from the given experimental conditions
Studying That Suits You
Use AI to generate personalized quizzes and flashcards to suit your learning preferences.
Related Documents
Description
Prepare for your BCH IS 3201 final exam with this comprehensive review covering essential lab math topics such as solution preparation, dilutions, and molarity calculations. You will also encounter practice problems related to making various concentrations and solutions. Test your understanding and ensure you're ready for the exam!
